Bacterial envelope, plasma membrane, and cell surface-associated proteins are the prime contact point of bacteria with their environment. Thus, the cell envelope composition and stress-induced changes are thought to be significantly related to the ability of bacteria to survive in a diverse environment. When bacterial cells are exposed to increased ethanol or butanol concentrations, some can survive and become ethanol or butanol tolerant. There are economic interests in studying bacterial ethanol tolerance particularly in wine making and in industrial ethanol fermentation. Strains that can tolerate higher ethanol or butanol concentrations are desired for industrial fermentation.
When bacterial cells are exposed to a high ethanol environment, changes to the outer envelopes and plasma membrane lipid bilayer have been reported (Ingram, L. O. 1976. Adaptation of membrane lipids to alcohols. J Bacteriol 125:670-8 and Malherbe, S., et al. 2012. Comparative metabolic profiling to investigate the contribution of O. oeni MLF starter cultures to red wine composition. J Ind Microbiol Biotechnol 39:477-94) and the protein expression profiles of both membrane proteins as well as cellular proteins were adjusted in response to ethanol (Liu, S. 2014. Proteomic analyses of ethanol tolerance in Lactobacillus buchneri NRRL B-30929. Proteomics 14:2540-4). Previous studies of bacterial isolates of the fuel ethanol production facilities suggested that species of lactic acid bacteria are commonly found. Changes in membrane lipid composition and fatty acid biosynthesis pathways have been shown by transcriptome analyses of Lactobacillus plantarum cells gown for 24 hours in 8% ethanol and changes of cytoplasmic and membrane proteins have also been reported in ethanol tolerant Oenococcus oeni cells grown in 8% ethanol (van Bokhorst-van de Veen, H., et al. Short- and long-term adaptation to ethanol stress and its cross-protective consequences in Lactobacillus plantarum. Appl Environ Microbiol 77:5247-56; and Silveira, M. G., et al. 2004. Effect of adaptation to ethanol on cytoplasmic and membrane protein profiles of Oenococcus oeni. Appl Environ Microbiol 70:2748-55). However, due to technical difficulties in obtaining a sufficient amount of membrane proteins, a comprehensive proteomics study of integral membrane proteins has not been possible.
Ethanol tolerance of Lactobacillus buchneri NRRL B-30929 strain has been first described in 2008 (Liu, S., et al. 2008. Lactobacillus buchneri strain NRRL B-30929 converts a concentrated mixture of xylose and glucose into ethanol and other products. J. Ind Microbiol Biotechnol 35:75-81). This strain L. buchneri NRRL B-30929, isolated from an ethanol production plant, was described as tolerant up to a 14% ethanol concentration in its growth environment. Further comparative 2D gel electrophoresis studies of proteins isolated from cells grown in media containing 10% ethanol vs. 0% ethanol revealed that a total of 308 protein spots were either up- or down-regulated by ethanol exposure. However, only 20 proteins (11 increased, 9 decreased) were identified in response to a 10% ethanol by proteomics analyses (Liu, S., et al. 2011. Complete genome sequence of Lactobacillus buchneri NRRL B-30929, a novel strain from a commercial ethanol plant. J. Bacteriol 193:4019-4020).
The traditional wine making process uses selected lactic acid bacteria to convert malic acid to lactic acid to reduce wine acidity after yeast alcoholic fermentation either via natural process or man-made inoculation. The malolactic fermentation (MLF) cannot only improve the wine stability, but can also enhance aroma, flavor, and taste attributed to the fermentative production of metabolic intermediates, and is therefore a desired and sought after trait.
O. oeni is regarded as one of the best malolactic bacteria that can survive the harsh winemaking conditions, especially the higher ethanol contents, and this species had therefore been the subject of numerous studies for its applications to enhance wine quality and aroma by producing a range of volatile aroma compounds such as diacetyl, acetoin and 2,3-butanediol (Alexandre, H., et al. 2004. Saccharomyces cerevisiae-Oenococcus oeni interactions in wine: current knowledge and perspectives. Int. J. Food Microbiol. 93:141-54 and Malherbe, 2012). Nevertheless ethanol tolerance is one of the essential criteria for developing malolactic fermentation strains to be introduced in the wine making process and further exploration regarding the ethanol tolerance trait of O. oeni is still important.
All of the references cited herein, including U.S. Patents and U.S. Patent Application Publications, are incorporated by reference in their entirety.
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